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Proteins Mar 2022Most mutations in the DNA-binding domain (DBD) of p53 inactivate or rescue the protein function interacting with the minor groove of DNA. However, how the conformation...
Most mutations in the DNA-binding domain (DBD) of p53 inactivate or rescue the protein function interacting with the minor groove of DNA. However, how the conformation changes propagating from the mutation sites result in distinct molecular recognition is still not well understood. As the protein mobility is an intrinsic property encrypted in its primary structure, we examined if different structures of wild-type and mutant p53 core domains display any unique patterns of intrinsic mobility. Normal mode calculation was employed to characterize the collective dynamics of DBD in p53 monomer and tetramer as well as their mutants. Intriguingly, the low-frequency collective motions of DBD show similar patterns between the wild-type protein and the rescued mutants. The analysis on atomic backbone fluctuations and low-frequency vibration mode statistics does further support the correlation between the intrinsic collective motion of DBD and the p53 protein function. The mutations in the DBD influence the low-frequency vibration of the p53 tetramer via the change of the collective motions among its four monomers. These findings thus provide new insights for understanding the physical mechanism of p53 protein structure-function relationship and help find the small molecule drug to modulate protein dynamic for disease therapy.
Topics: Amino Acid Sequence; Binding Sites; DNA; Humans; Models, Molecular; Mutant Proteins; Mutation; Protein Binding; Protein Domains; Structure-Activity Relationship; Tumor Suppressor Protein p53
PubMed: 34792219
DOI: 10.1002/prot.26283 -
Biochemistry Jan 2021Rhodopsin is the light receptor required for the function and health of photoreceptor cells. Mutations in rhodopsin can cause misfolding and aggregation of the receptor,...
Rhodopsin is the light receptor required for the function and health of photoreceptor cells. Mutations in rhodopsin can cause misfolding and aggregation of the receptor, which leads to retinal degeneration. Bovine rhodopsin is often used as a model to understand the effect of pathogenic mutations in rhodopsin due to the abundance of structural information on the bovine form of the receptor. It is unclear whether or not the bovine rhodopsin template is adequate in predicting the effect of these mutations occurring in human retinal disease or in predicting the efficacy of therapeutic strategies. To better understand the extent to which bovine rhodopsin can serve as a model, human and bovine P23H rhodopsin mutants expressed heterologously in cells were examined. The aggregation properties and cellular localization of the mutant receptors were determined by Förster resonance energy transfer and confocal microscopy. The potential therapeutic effects of the pharmacological compounds 9- retinal and metformin were also examined. Human and bovine P23H rhodopsin mutants exhibited different aggregation properties and responses to the pharmacological compounds tested. These observations would lead to different predictions on the severity of the phenotype and divergent predictions on the benefit of the therapeutic compounds tested. The bovine rhodopsin template does not appear to adequately model the effects of the P23H mutation in the human form of the receptor.
Topics: Animals; Cattle; Diterpenes; Humans; Metformin; Mutant Proteins; Mutation; Protein Aggregates; Retinaldehyde; Rhodopsin
PubMed: 33356167
DOI: 10.1021/acs.biochem.0c00733 -
Biomolecules Nov 2020The bacterial RNA polymerase (RNAP) is a multi-subunit protein complex (α2ββ'ω σ) containing the smallest subunit, ω. Although identified early in RNAP research,... (Review)
Review
The bacterial RNA polymerase (RNAP) is a multi-subunit protein complex (α2ββ'ω σ) containing the smallest subunit, ω. Although identified early in RNAP research, its function remained ambiguous and shrouded with controversy for a considerable period. It was shown before that the protein has a structural role in maintaining the conformation of the largest subunit, β', and its recruitment in the enzyme assembly. Despite evolutionary conservation of ω and its role in the assembly of RNAP, mutants lacking (codes for ω) are viable due to the association of the global chaperone protein GroEL with RNAP. To get a better insight into the structure and functional role of ω during transcription, several dominant lethal mutants of ω were isolated. The mutants showed higher binding affinity compared to that of native ω to the α2ββ' subassembly. We observed that the interaction between α2ββ' and these lethal mutants is driven by mostly favorable enthalpy and a small but unfavorable negative entropy term. However, during the isolation of these mutants we isolated a silent mutant serendipitously, which showed a lethal phenotype. Silent mutant of a given protein is defined as a protein having the same sequence of amino acids as that of wild type but having mutation in the gene with alteration in base sequence from more frequent code to less frequent one due to codon degeneracy. Eventually, many silent mutants were generated to understand the role of rare codons at various positions in . We observed that the dominant lethal mutants of ω having either point mutation or silent in nature are more structured in comparison to the native ω. However, the silent code's position in the reading frame of plays a role in the structural alteration of the translated protein. This structural alteration in ω makes it more rigid, which affects the plasticity of the interacting domain formed by ω and α2ββ'. Here, we attempted to describe how the conformational flexibility of the ω helps in maintaining the plasticity of the active site of RNA polymerase. The dominant lethal mutant of ω has a suppressor mapped near the catalytic center of the β' subunit, and it is the same for both types of mutants.
Topics: Bacterial Proteins; DNA-Directed RNA Polymerases; Mutant Proteins; Protein Subunits; Structure-Activity Relationship; Transcription Factors
PubMed: 33238579
DOI: 10.3390/biom10111588 -
Journal of Molecular Biology Aug 2017Influenza virus evolves rapidly to constantly escape from natural immunity. Most humoral immune responses to influenza virus target the hemagglutinin (HA) glycoprotein,... (Review)
Review
Influenza virus evolves rapidly to constantly escape from natural immunity. Most humoral immune responses to influenza virus target the hemagglutinin (HA) glycoprotein, which is the major antigen on the surface of the virus. The HA is composed of a globular head domain for receptor binding and a stem domain for membrane fusion. The major antigenic sites of HA are located in the globular head subdomain, which is highly tolerant of amino acid substitutions and continual addition of glycosylation sites. Nonetheless, the evolution of the receptor-binding site and the stem region on HA is severely constrained by their functional roles in engaging the host receptor and in mediating membrane fusion, respectively. Here, we review how broadly neutralizing antibodies (bnAbs) exploit these evolutionary constraints to protect against diverse influenza strains. We also discuss the emerging role of other epitopes that are conserved only in subsets of viruses. This rapidly increasing knowledge of the evolutionary biology, immunology, structural biology, and virology of influenza virus is invaluable for development and design of more universal influenza vaccines and novel therapeutics.
Topics: Antibodies, Neutralizing; Antibodies, Viral; Evolution, Molecular; Hemagglutinin Glycoproteins, Influenza Virus; Immune Evasion; Mutant Proteins; Mutation; Orthomyxoviridae
PubMed: 28648617
DOI: 10.1016/j.jmb.2017.06.015 -
Endocrine Nov 2018The importance of androgen receptor variants (AR-Vs) is recognized in prostate cancer. AR-Vs have been the focus of many studies. Expression of AR-Vs has been proposed...
PURPOSE
The importance of androgen receptor variants (AR-Vs) is recognized in prostate cancer. AR-Vs have been the focus of many studies. Expression of AR-Vs has been proposed as a biomarker for resistance to androgen deprivation therapy for metastatic disease. Herein, we show dynamic changes in AR-Vs expression in response to androgen modulation.
METHODS
The C4-2B cell line was exposed to low (10 M) and high (10 M) androgen (dihydrotestosterone, DHT) levels, with or without flutamide. mRNA and protein expression levels were assessed by qPCR and immunohistochemistry, respectively.
RESULTS
We demonstrated that high levels of DHT downregulate AR-FL and AR-Vs. Even though AR-Vs did not present ligand-binding domain, thus were not capable of binding to DHT, they present dynamic changes under androgen treatment. Treatment with flutamide alone or in association with low levels of DHT stimulates growth of prostatic cells.
CONCLUSIONS
Importantly, we provide evidence that AR-Vs respond differently to androgenic modulation. These findings have implications for a better understanding of the role of AR-Vs in prostate carcinogenesis.
Topics: Androgens; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; Humans; Ligands; Male; Mutant Proteins; Polymorphism, Genetic; Prostatic Neoplasms; Protein Isoforms; Receptors, Androgen
PubMed: 30027434
DOI: 10.1007/s12020-018-1682-5 -
Cells Nov 2021The wild-type protein p53 plays a key role in preventing the formation of neoplasms by controlling cell growth. However, in more than a half of all cancers, the gene... (Review)
Review
The wild-type protein p53 plays a key role in preventing the formation of neoplasms by controlling cell growth. However, in more than a half of all cancers, the gene has missense mutations that appear during tumorigenesis. In most cases, the mutated gene encodes a full-length protein with the substitution of a single amino acid, resulting in structural and functional changes and acquiring an oncogenic role. This dual role of the wild-type protein and the mutated isoforms is also evident in the regulation of the redox state of the cell, with antioxidant and prooxidant functions, respectively. In this review, we introduce a new concept of the p53 protein by discussing its sensitivity to the cellular redox state. In particular, we focus on the discussion of structural and functional changes following post-translational modifications of redox-sensitive cysteine residues, which are also responsible for interacting with zinc ions for proper structural folding. We will also discuss therapeutic opportunities using small molecules targeting cysteines capable of modifying the structure and function of the p53 mutant isoforms in view of possible anticancer therapies for patients possessing the mutation in the gene.
Topics: Animals; Cysteine; Humans; Mutant Proteins; Oxidation-Reduction; Protein Isoforms; Protein Processing, Post-Translational; Structure-Activity Relationship; Tumor Suppressor Protein p53
PubMed: 34831372
DOI: 10.3390/cells10113149 -
Cell Death & Disease May 2021The nucleus is the target of autoantibodies in many diseases, which suggests intrinsic nuclear adjuvants that confer its high autoimmunogenicity. Nucleolin (NCL) is one...
The nucleus is the target of autoantibodies in many diseases, which suggests intrinsic nuclear adjuvants that confer its high autoimmunogenicity. Nucleolin (NCL) is one abundant nucleolar autoantigen in systemic lupus erythematosus (SLE) patients and, in lupus-prone mice, it elicits autoantibodies early. With purified NCL, we observed that it was a potent alarmin that activated monocytes, macrophages and dendritic cells and it was a ligand for TLR2 and TLR4. NCL released by necrotic cells also exhibited alarmin activity. The NCL alarmin activity resides in its glycine/arginine-rich (GAR/RGG) motif and can be displayed by synthetic GAR/RGG peptides. Two more GAR/RGG-containing nucleolar proteins, fibrillarin (FBRL) and GAR1, were also confirmed to be novel alarmins. Therefore, the GAR/RGG alarmin motif predicts a family of nucleolar alarmins. The apparent prevalence of nucleolar alarmins suggests their positive contribution to tissue homeostasis by inducing self-limiting tissue inflammation with autoimmunity only occurring when surveillance is broken down.
Topics: Alarmins; Animals; Cell Nucleus; Humans; Mice; Mutant Proteins
PubMed: 33980825
DOI: 10.1038/s41419-021-03766-w -
Mini Reviews in Medicinal Chemistry 2016Isocitrate dehydrogenase (IDH) is a metabolic enzyme that converts isocitrate to α-ketoglutarate (α-KG). Genetic gain-of-function mutations in IDH1 and IDH2 confer a... (Review)
Review
Isocitrate dehydrogenase (IDH) is a metabolic enzyme that converts isocitrate to α-ketoglutarate (α-KG). Genetic gain-of-function mutations in IDH1 and IDH2 confer a neomorphic activity that allow reduction of α -KG to (R)-2- hydroxyglutarate, the accumulation of which results in the development of cancers like low grade gliomas and leukemia. After treatment with AG-221 in clinical trials, a first-in-class inhibitor of mutated IDH2, 29 patients with acute myeloid leukemia or myelodysplastic syndrome experience complete remissions and the overall response rate is 59/159 (37%). Thus, IDH mutants have become intriguing targets for cancer therapeutics. In addition to providing a brief summary of IDH mutations, this review describes known inhibitors with potential activities against IDH mutants such as AG-120, AG-221, AG-881 and AGI-6780. The evolving landscape of IDH mutant inhibitors provides us an outlook on the discovery of novel, safer, and more effective cancer treatment strategies.
Topics: Enzyme Inhibitors; Humans; Isocitrate Dehydrogenase; Mutant Proteins; Mutation; Neoplasms
PubMed: 27292784
DOI: 10.2174/1389557516666160609085520 -
Molecular Microbiology Mar 2019Tail-anchored (TA) proteins are membrane proteins that are found in all domains of life. They consist of an N-terminal domain that performs various functions and a...
Tail-anchored (TA) proteins are membrane proteins that are found in all domains of life. They consist of an N-terminal domain that performs various functions and a single transmembrane domain (TMD) near the C-terminus. In eukaryotes, TA proteins are targeted to the membranes of mitochondria, the endoplasmic reticulum (ER), peroxisomes and in plants, chloroplasts. The targeting of these proteins to their specific destinations correlates with the properties of the C-terminal domain, mainly the TMD hydrophobicity and the net charge of the flanking regions. Trichomonas vaginalis is a human parasite that has adapted to oxygen-poor environment. This adaptation is reflected by the presence of highly modified mitochondria (hydrogenosomes) and the absence of peroxisomes. The proteome of hydrogenosomes is considerably reduced; however, our bioinformatic analysis predicted 120 putative hydrogenosomal TA proteins. Seven proteins were selected to prove their localization. The elimination of the net positive charge in the C-tail of the hydrogenosomal TA4 protein resulted in its dual localization to hydrogenosomes and the ER, causing changes in ER morphology. Domain mutation and swap experiments with hydrogenosomal (TA4) and ER (TAPDI) proteins indicated that the general principles for specific targeting are conserved across eukaryotic lineages, including T. vaginalis; however, there are also significant lineage-specific differences.
Topics: DNA Mutational Analysis; Multienzyme Complexes; Mutant Proteins; Organelles; Protein Transport; Protozoan Proteins; Recombinant Proteins; Trichomonas vaginalis
PubMed: 30506591
DOI: 10.1111/mmi.14175 -
Journal of Molecular Biology Jun 2019Signal recognition particle (SRP) recognizes signal sequences of secretory proteins and targets them to the endoplasmic reticulum membrane for translocation. Many human...
Signal recognition particle (SRP) recognizes signal sequences of secretory proteins and targets them to the endoplasmic reticulum membrane for translocation. Many human diseases are connected with defects in signal sequences. The current dogma states that the molecular basis of the disease-associated mutations in the secretory proteins is connected with defects in their transport. Here, we demonstrate for several secretory proteins with disease-associated mutations that the molecular mechanism is different from the dogma. Positively charged or helix-breaking mutations in the signal sequence hydrophobic core prevent synthesis of the aberrant proteins and lead to degradation of their mRNAs. The degree of mRNA depletion depends on the location and severity of the mutation in the signal sequence and correlates with inhibition of SRP interaction. Thus, SRP protects secretory protein mRNAs from degradation. The data demonstrate that if disease-associated mutations obstruct SRP interaction, they lead to silencing of the mutated protein expression.
Topics: Disease; HeLa Cells; Humans; Mutant Proteins; Mutation; Proteins; RNA, Messenger; Signal Recognition Particle
PubMed: 31100385
DOI: 10.1016/j.jmb.2019.05.011